The Mekong River has the largest inland fishery in the world but is threatened by planned mainstem hydropower developments that could impact fish migration patterns. Here we provide direct evidence of long-distance migrations along the Mekong mainstem by Hypsibarbus malcomi, one of the 10 most important species in the Mekong River fishery. The migration included a 354 km passage through one current and three proposed hydropower developments. We tagged and released 233 wild fish, including 77 H. malcomi with Passive Integrated Transponder (PIT tags) near the capital Vientiane in April of 2022. Five PIT tagged H. malcolmi were detected at the top of the Xayaburi hydropower plant fish ladder within 15 months of being released at Vientiane. Our findings empirically demonstrate that this species undertakes long distance migrations in the main stem of the Mekong and therefore that suitable fish passage facilities will need to be incorporated in mainstem hydropower developments to meet their migratory needs. The findings also demonstrate the value of including a fishery independent and efficient monitoring technology, such as PIT tagging, to assess the migration patterns of fish in the face of upcoming developments in the Lower Mekong Basin.